Shell Vial Assay Research Articles

Comparison of cytomegalovirus antigenemia and culture assays in patients on and off antiviral therapy

We examined 1,869 consecutive blood specimens from 529 patients (>80% organ transplant recipients) for detection of CMV by antigenemia and culture assays, and compared results between patients on and off antiviral therapy. All 1,869 specimens were tested by the shell vial assay and antigenemia, and 503 were also tested by standard tube culture. The overall positivity rate for each test was 17.0% for antigenemia, 1.8% for shell vial culture assay, and 0.7% by tube culture. No specimens were positive by either shell vial or tube culture, while negative by antigenemia. These findings were consistent across all organ transplant and other patient types. Shell vial positivity was associated with higher antigenemia levels in patients either on or off anti-CMV drug therapy. Among the shell vial positive specimens, the antigenemia counts were higher in patients on antiviral drug therapy as compared to those not on therapy. We conclude that the pp65 antigenemia assay is superior to culture methods for detection of CMV in blood, particularly for patients on anti-CMV drug treatment. Additionally, its quantitative nature renders the antigenemia assay an excellent tracking tool for both resolution of asymptomatic, low level CMV reactivations and response of CMV infection to antiviral treatment. J. Med. Virol. 59:91–97, 1999. © 1999 Wiley-Liss, Inc.

Use of molecular assays in diagnosis and monitoring of cytomegalovirus disease following renal transplantation.

We compared two commercial molecular assays (the Murex Hybrid Capture CMV DNA assay [HCA], version 2, and the Roche Amplicor plasma PCR assay) with a standard shell vial assay in detecting and predicting cytomegalovirus (CMV) disease in a group of renal transplant patients and assessed the role of viral load measurements (using the HCA) in their management. The sensitivity of the HCA and Amplicor assay in terms of disease detection was 100%, compared to 71% for the shell vial assay. Both the HCA and the PCR assay detected all cases of disease, at medians of 11 and 12.5 days before the onset of symptoms, respectively. Significantly higher viral loads were detected in those patients with symptoms (7.9 x 10(5) copies/ml) than in patients without symptoms (7.9 x 10(4) copies/ml; P < 0.0001). There was also a trend towards higher viral loads in those patients with primary infections (7.8 x 10(5) copies/ml) than in those patients with reactivations of CMV disease or reinfections. Successful treatment with ganciclovir was associated with a >90% reduction in viral load. Both of these new assays are sensitive and easy to use. A comparison of accurate quantitation is also useful in monitoring responses to antiviral therapy.

The development of an automated in situ assay for the detection of human cytomegalovirus in peripheral blood leukocytes

Human cytomegalovirus (HCMV) infections are common in immunosuppressed patients, especially transplant recipients and patients with AIDS. The utility of an automated in situ hybridization (ISH) assay for the rapid detection of HCMV immediate early mRNA was evaluated using cytospin (Shandon Lipshaw, Inc., Pittsburgh, PA) prepared leukocytes from peripheral blood samples. In this study, the detection of HCMV immediate early protein by immunofluorescent antibody staining of the standard shell vial assay was compared to the detection of HCMV immediate early mRNA in peripheral blood leukocytes using the automated ISH system. Of 135 specimens tested, eight specimens were positive using HCMV ISH compared to seven positive specimens using shell vial assay. Overall, HCMV ISH demonstrated 100% sensitivity and 99% specificity. Since the HCMV ISH assay requires minimal labor, and can be completed in less than 5 h, this method should be evaluated as a potential replacement for shell vial assay for the diagnosis of HCMV infection.

Rapid detection of respiratory viruses by shell vial assay using simultaneous culture of HEp-2, LLC-MK2, and MDCK cells in a single vial.

A shell vial assay with simultaneous culture of HEp-2, LLC-MK2, and MDCK cell lines in a single tube (CoHLM SV assay) was compared with traditional tube culture (TC) for the detection of the main respiratory viruses in 358 nasal wash specimens. A total of 170 strains were isolated from 168 virus-positive samples. A total of 94. 1% of the strains (160 strains; 128 respiratory syncytial viruses and 32 other viruses) were detected by the CoHLM SV assay in 48 h, whereas 98.2% of the strains (167 strains; 132 respiratory syncytial viruses and 35 other viruses) were detected by TC in a mean time of 6 days. The CoHLM SV assay may be useful for the rapid detection of respiratory viruses.

Culture of Bartonella quintana and Bartonella henselae from human samples: a 5-year experience (1993 to 1998).

Bartonella quintana and Bartonella henselae are fastidious gram-negative bacteria responsible for bacillary angiomatosis, trench fever, cat scratch disease, and endocarditis. During a 5-year period, we received 2,043 samples for culture of Bartonella sp. We found Bartonella sp. to be the etiologic agent in 38 cases of endocarditis, 78 cases of cat scratch disease, 16 cases of bacteremia in homeless people, and 7 cases of bacillary angiomatosis. We correlated the results of positive cultures with the clinical form of the disease, type of sample, culture procedure, PCR-based genomic detection, and antibody determination. Seventy-two isolates of B. quintana and nine isolates of B. henselae from 43 patients were obtained. Sixty-three of the B. quintana isolates and two of the B. henselae isolates, obtained from patients with no prior antibiotic therapy, were stably subcultured. The sensitivity of culture was low when compared with that of PCR-based detection methods in valves of patients with endocarditis (44 and 81%, respectively), skin biopsy samples of patients with bacillary angiomatosis (43 and 100%, respectively), and lymph nodes of cat scratch disease (13 and 30%, respectively). Serological diagnosis was also more sensitive in cases of endocarditis (97%) and cat scratch disease (90%). Among endocarditis patients, the sensitivity of the shell vial culture assay was 28% when inoculated with blood samples and 44% when inoculated with valvular biopsy samples, and the sensitivity of both was significantly higher than that of culture on agar (5% for blood [P = 0.045] and 4% for valve biopsy samples [P < 0.0005]). The most efficient culture procedure was the subculture of blood culture broth into shell vials (sensitivity, 71%). For patients with endocarditis, previous antibiotic therapy significantly affected results of blood culture; no patient who had been administered antibiotics yielded a positive blood culture, whereas 80% of patients with no previous antibiotic therapy yielded positive blood cultures (P = 0.0006). Previous antibiotic therapy did not, however, prevent isolation of Bartonella sp. from cardiac valves but did prevent the establishment of strains, as none of the 15 isolates from treated patients could be successfully subcultured. For the diagnosis of B. quintana bacteremia in homeless people, the efficiency of systematic subculture of blood culture broth onto agar was higher than that of direct blood plating (respective sensitivities, 98 and 10% [P < 10(-7)]). Nevertheless, both procedures are complementary, since when used together their sensitivity reached 100%. All homeless people with positive blood cultures had negative serology. The isolation rate of B. henselae from PCR-positive lymph nodes, in patients with cat scratch disease, was significantly lower than that from valves of endocarditis patients and skin biopsy samples from bacillary angiomatosis patients (13 and 33%, respectively [P = 0.084]). In cases of bacillary angiomatosis for which an agent was identified to species level, the isolation rate of B. henselae was lower than the isolation rate of B. quintana (28 and 64%, respectively [P = 0.003]). If culture is to be considered an efficient tool for the diagnosis of several Bartonella-related diseases, methodologies need to be improved, notably for the recovery of B. henselae from lymph nodes of patients with cat scratch disease.

Longitudinal Analysis of Plasma Cytomegalovirus DNA in a Child with Crohn's Disease and Cytomegalovirus Gastroenteritis

Longitudinal Analysis of Plasma Cytomegalovirus DNA in a Child with Crohn's Disease and Cytomegalovirus Gastroenteritis

Detection of mumps virus in clinical specimens by rapid centrifugation culture and conventional tube cell culture

Conventional tube cell culture was compared with a 2 day and 5 day spin-amplified shell vial indirect immunofluorescence assay for the detection of mumps virus in swabs from the area of Stensen's duct. The sensitivity and specificity of the shell vial assay were 95.9 and 100%, respectively. The shell vial detected 66.3% of the positive cultures within 2 days of inoculation while the first positive results were available by conventional tube cell culture after 3 days (1.6%) reaching 72.4% of all culture positive specimens after 7 days. These data suggest that a centrifugation shell vial indirect immunofluorescence assay may be useful for rapid detection of mumps virus in clinical specimens.

Application of polymerase chain reaction to the detection of cytomegalovirus in bronchoalveolar lavage from lung transplant recipients

The use of polymerase chain reaction (PCR) for the detection of cytomegalovirus (CMV) in bronchoalveolar lavages (BAL) of immunocompromised patients was evaluated. Firstly, PCR was compared to conventional cell culture (CCC) and shell vial assay (SVA) in 65 BAL received in our laboratory on a routine basis. Sensitivity and specificity of the PCR were 100% and 87.2% respectively. The PCR method was then applied to the monitoring of 55 BAL from nine lung transplant recipients. The data illustrate the excellent sensitivity of PCR. However the predictive value of the test for the occurrence of a symptomatic infection was weak.

Evaluation of 2-SP transport medium for detection of Chlamydia trachomatis and Neisseria gonorrhoeae by two automated amplification systems and culture for chlamydia.

AIMS: To assess the performance of 2-sucrose-phosphate based transport medium (2-SP) for the detection of Chlamydia trachomatis and Neisseria gonorrhoeae by an automated commercial polymerase chain reaction (PCR) and ligase...

Monitoring and diagnosis of cytomegalovirus infection in renal transplantation.

In this study, the utility of the cytomegalovirus antigen (CMV-AG) and the shell vial (SV) tests in the diagnosis and monitoring of posttransplant CMV infection were compared. Previous retrospective studies from the authors' center suggested that the CMV-AG test, which uses monoclonal antibodies to detect viral antigen in circulating peripheral blood leukocytes (PBL) may be both a more sensitive and specific test. A cohort of 32 renal transplant recipients was followed-up prospectively with serial CMV-AG testing, as well as conventional culture and SV for blood and urine and tests for immunoglobulin M (IgM) antibody. It was discovered that the CMV-AG test was not only more sensitive than the SV test in detecting CMV infection, but that the degree of antigenemia as expressed by the number of positive cells per 50,000 PBL correlated with the likelihood and degree of symptomatic infection. All patients with a count > 10 positive cells/50,000 PBL developed clinical symptoms; therefore, this threshold could be useful in deciding clinically whether fever is related to CMV infection. Alternatively, if antigenemia were monitored serially after transplant, the same threshold could be used as a trigger for instituting antiviral therapy, because it was often reached prior to the onset of symptoms and had a high specificity for subsequent symptomatic infection. Such an approach could obviate unnecessary treatment of patients not destined to become symptomatic. Based upon the findings in this study, the CMV-AG test is superior to the SV assay because the actual count helps determine the likelihood that symptoms are a result of the virus and the processing time is shorter, it can be used to monitor the response to therapy and as a guide to the institution of preemptive therapy.

Significance of leukocyte concentration in the performance of the quantitative cytomegalovirus (CMV) antigenemia assay

Background: The quantitative cytomegalovirus-antigenemia (CMV-Ag) assay is an important technology in the regimen of tests utilized in the care and management of acquired immunodeficiency syndrome (AIDS) and other immunocompromised patient groups. Performance of this assay is contingent upon the appropriate processing of the polymorphonuclear leukocyte (PMNL) compartment of the peripheral blood. However, a cell input standard in the performance of the CMV-Ag assay, has not been established. Interpretive differences between laboratories utilizing the CMV-Ag assay may reflect this lack of test uniformity. Objectives: To determine the effect of different PMNL concentrations on the quantitation of CMV in peripheral blood. The leukocyte concentration resulting in optimal rates of viral detection, will be compared with the shell vial assay-indirect immunofluorescent assay (SVA-IFA), and conventional tube culture (TC-CPE). Study design: A total of 74 freshly collected blood specimens were tested by the CMV-Ag assay, using cytospin preparations consisting of 2×10 5, 4×10 5 and in preliminary experiments, 8×10 5 PMNLs/slide. Data obtained from these studies were compared to SVA-IFA and TC-CPE. Viral load was monitored among 11 symptomatic patients through sequential testing of these patients at the start of ganciclovir (GCV), foscarnet (PFA), or combination drug therapy. Results: Among 74 blood specimens tested by the CMV-Ag assay, cytospin preparations consisting of 4×10 5 compared with 2×10 5 PMNLs/slide, affected a mean positive cell increase of 215% ( P=0.03). PMNL slide preparations consisting 8×10 5 cells produced background levels which prevented accurate reading of slides. The CMV-Ag assay was more sensitive than the SVA-IFA, but equivalent to TC-CPE. Among 11 patients started on drug therapy, viral load was markedly reduced in 8 within 2–3 weeks; three patients (2 deceased within 3 weeks after receiving therapy), showed no decrease in viral load. One patient was identified as harboring a PFA resistant strain. Conclusions: A PMNL concentration of 4×10 5 cells facilitated the reading of CMV-Ag assay slide preparations. The modified CMV-Ag assay furthermore, is applicable in the monitoring of viral load for the tracking of susceptible or resistant CMV strains.

Bacteriostatic and bactericidal activity of levofloxacin against Rickettsia rickettsii, Rickettsia conorii, 'Israeli spotted fever group rickettsia' and Coxiella burnetii.

Levofloxacin, the L-isomer of ofloxacin, is approximately twice as active as ofloxacin against most Gram-positive and Gram-negative bacteria, and has improved intracellular pharmacokinetic and pharmacodynamic properties. The present work deals with the in-vitro activity of levofloxacin against the obligate intracellular bacteria Rickettsia rickettsii, Rickettsia conorii, 'Israeli spotted fever group rickettsia' (Israeli SFGR) and Coxiella burnetii. Fluoroquinolones, including ofloxacin, have previously been shown to be bacteriostatic against Rickettsia spp. and C. burnetii in vitro. They are reliable alternatives to tetracycline therapy for Mediterranean spotted fever, scrub typhus and acute Q fever. Levofloxacin was bacteriostatic against R. rickettsii, R. conorii and the Israeli SFGR at concentrations of 0.5-1 mg/L, as determined by both a plaque assay and a dye uptake assay. It was also bacteriostatic against C. burnetii isolates, including the Nine Mile, Priscilla and Q212 strains, at concentrations of 0.5-2 mg/L, as determined using the shell vial assay. Overall, levofloxacin could inhibit rickettsial growth at concentrations equal to or half of those necessary for growth inhibition by ofloxacin. Levofloxacin was not bactericidal against C. burnetii at concentrations up to 4 mg/L.

USEFULNESS OF DNA VIRAL LOAD QUANTIFICATION FOR CYTOMEGALOVIRUS DISEASE MONITORING IN RENAL AND PANCREAS/RENAL TRANSPLANT RECIPIENTS1

The purpose of this prospective study was to evaluate the usefulness of quantifying DNA-cytomegalovirus (CMV) load for the diagnosis and monitoring of CMV disease among renal and pancreas transplant patients under immunosuppressive drugs. A longitudinal study was conducted among 34 consecutive, unselected renal and pancreas/renal transplanted patients in our unit. During the first 3 posttransplant months, weekly monitoring of CMV infection and CMV disease was done, involving the determination of viremia by the shell vial assay, qualitative DNAemia by semi-nested polymerase chain reaction (PCR) and quantitative DNAemia by the hybrid capture system (HCS), a new and original hybridization method (337 samples were collected for each test). Qualitative and quantitative DNAemia results were blinded to physicians and three grades of disease were defined according to CMV related symptom occurrence. PCR was the most sensitive (100%) but the least specific (78%) method for the diagnosis of CMV disease. HCS was specific for CMV genome detection, sensitive and reproducible. Blood DNA levels above 60 pg/ml were predictive of severe or moderate CMV disease (sensitivity, 92%; specificity, 100%). A significant decrease in viral load was observed after ganciclovir administration, and a positive PCR or HCS result at the end of the antiviral treatment was associated with relapse of CMV infection or disease. It is concluded that quantitative DNAemia detection, with this new commercially available method, can predict disease and may be useful for a rational evaluation of ganciclovir preemptive therapy in such patients.

Determination of the number of blood samples needed for optimal detection of cytomegalovirus viremia in immunocompromised patients using a shell-vial assay.

To establish the number of blood samples necessary for the diagnosis of viremic episodes caused by cytomegalovirus (CMV), a prospective analysis was conducted of 238 patients (38 renal transplant recipients and 200 HIV-infected patients) who developed CMV viremia. The usefulness of samples and the volume of blood required to demonstrate the presence of viremia by CMV was also studied. The first blood sample was diagnostic for CMV viremia in 53.3% of the viremic patients; the second sample documented an additional 22.2% of cases of viremia (75.5% of infected patients); and the third sample demonstrated viremia in the remaining 24.5%. Thus, a diagnosis of CMV viremia was established in every patient (100% of episodes of viremia). In this study, the use of three 3 ml blood samples collected at 24 h intervals was sufficient to detect all episodes of CMV viremia in patients clinically suspected to have disseminated disease.

Use of DNA amplification for diagnosis of cytomegalovirus enteritis after intestinal transplantation

BACKGROUND & AIMS: Intestinal transplantation is a feasible therapy for patients with short-bowel syndrome. However, cytomegalovirus (CMV) enteritis can cause complications. The aim of this study was to investigate the value of polymerase chain reaction (PCR)-based detection methods for CMV in the management of patients with small bowel transplants. METHODS: Comparative evaluation of PCR with histopathology, shell-vial assay, and tube culture of intestinal biopsy specimens was used for the diagnosis of CMV enteritis in 21 patients. RESULTS: Ten patients experienced 21 episodes of CMV enteritis, diagnosed by histopathology, virology, or both. PCR had a sensitivity and specificity of 96% and 69%, respectively, compared with traditional methods, whereas the positive and negative predictive values were 35% and 99%, respectively. Three+ and 4+ signals corresponded to a specificity of 91% and positive predictive value of 59%, respectively. CMV was detected by PCR a median of 11 days (range, 0-32) earlier than other methods and lasted a median of 40 days (range, 21-80) in the 13 episodes, which became PCR-negative and in those patients who developed asymptomatic infection. In 8 episodes, CMV by PCR never became negative and was associated with a relapse of disease confirmed by other methods. CONCLUSIONS: PCR is a sensitive method for the early detection of CMV in intestinal biopsy specimens and can be used for preemptive therapy after intestinal transplantation. (Gastroenterology 1997 Apr;112(4):1121-8)

A rapid culture assay for examining measles virus infections from urine specimens

Background: Large numbers of measles virus (MV) specimens are processed in our laboratory each year as part of a molecular epidemiological study of MV in South Africa. The development of a sensitive, rapid virus isolation system is needed to cope with the number of specimens processed. Objectives: A comparison was made of centrifugation-enhanced shell vial culture and standard tissue culture using B95a cells for the isolation of MV from throat swabs and urine. Study design: The rapid method was initially evaluated using Schwarz vaccine virus and then compared to standard culture using throat swab specimens. Results: The shell vial assay proved to be ten times more sensitive than standard culture in the initial evaluation. Of 43 throat swab specimens, 37 (86%) were positive and 6 (14%) negative in standard culture using B95a cells. The specimens were removed after adsorption in standard culture, frozen and then used in the shell vial assay. It was found that 16 27 were positive in the shell vial assay (24 of these 27 being positive in standard culture,) and 8 negative and 8 specimens gave an indeterminate result. For the 45 urine specimens used in the shell vial assay, 71% were positive, 11% negative and 18% gave an indeterminate result, due to too few cells being present for antigen determination by indirect fluorescent antibody assay. Results were obtained in 4 days, as opposed to the average of 14 days for confirmed isolation in standard culture. Conclusion: Rapid culture substantially reduced total test time, was less labour-intensive and was as sensitive as standard culture for the isolation of measles virus from clinical specimens.

Comparison of cytomegalovirus antigenemia and shell vial culture in allogeneic marrow transplantation recipients receiving ganciclovir prophylaxis.

We prospectively monitored 61 allogeneic BMT patients for evidence of CMV infection and disease starting 7 days prior to transplant until day 110 after transplant. Patients receiving pre- and post-transplantation ganciclovir prophylaxis were followed for the incidence of infection by the CMV antigenemia assay and shell vial cultures. The median age of all patients was 32 years (range 5-54 years). Fourteen (25%) of 57 evaluable patients became CMV antigenemia or culture positive. The incidence of culture or antigenemia positivity in CMV seropositive or seronegative patients with a seropositive donor was 29% (14 of 49 patients). The antigenemia assay became positive a median of 29 days (range 12-89 days) after BMT as compared to 46 days (range 26-98 days) by shell vial assay (P < 0.001). There were no cases of CMV disease in the first 110 days after transplant. This study demonstrates that despite the use of prophylactic ganciclovir, BMT patients developed CMV infection but did not progress to disease in this study, the CMV antigenemia assay may be used to monitor for CMV infection during prophylaxis, and the current regimens for CMV prophylaxis with ganciclovir may require further evaluation to determine an optimal regimen to prevent CMV infection.

The laboratory diagnosis of cytomegalovirus infections

Rapid and accurate diagnosis of cytomegalovirus (CMV) infection is imperative with the advent of effective antiviral therapy (gangiclovir, foscarnet, CMV hyperimmune globin). Applications of conventional cell culture (CC), shell vial assay (SV), serological testing, antigenemia assay (AG) as well as molecular methods [polymerase chain reaction (PCR), branch DNA (b-DNA) and hybrid capture (HC)] to various patient populations and specimen types are discussed. A three year study of 670 specimens [354 urines, 205 peripheral blood leukocytes (PBLs), 56 upper respiratory and 55 tissues] compared CMV CC and SV isolation rates. Of the total, 124 (18.5%) were positive by either or both techniques. For each specimen type the number of positives detected by SV was greater than CC (urine 28 vs 15, PBLs, 12 vs 2). However, of 124 positives, 21 were solely CC positive. A comparison of SV to AG in 230 PBLs yielded a sensitivity of 100% and specificity of 68.3%. The low specificity when compared to SV may be due to the increased sensitivity of AG. Fifty-nine PBLs were examined for differing immunostaining techniques [Immunoperoxidase (IP) vs Immunofluorescence (IF)]. IF stained PBLs showed an increased number of positive cells per preparation and greater stain intensity for ease of interpretation.

Polymerase Chain Reaction Assays for the Detection of Cytomegalovirus in Organ and Bone Marrow Transplant Recipients

Cytomegalovirus (CMV) infection is ubiquitous and results in a wide spectrum of clinical manifestations ranging from asymptomatic infection to severe life threatening disease. Infection in normal children and adults usually causes no symptoms but in the immuncompromised host, CMV may result in severe opportunistic infections with high morbidity and mortality. Historically, virus detection was dependent on culture of the virus or on a centrifugation culture system referred to as a shell vial assay. The shell vial assay frequently lacked sensitivity and was unable to detect infection in its early phase. Also, as with culture assays, the results were affected by antiviral therapy. The CMV antigenemia assay was developed to provide more rapid results and has gained wide usage. This assay is limited to detection of the virus in white blood cells and is more sensitive than culture or the shell vial assay.Application of the polymerase chain reaction (PCR) to these problems has resulted in the development of assays for CMV which are more sensitive than previously available methods. This method employs liquid hybridization with 32P labeled probes and gel retardation analysis for detection of amplified DNA specific for each virus.A comparison of the detection of CMV by an antigenemia assay or the PCR method in the leukocytes of renal transplant patients revealed that the PCR assay detects cytomegalovirus earlier and more consistently than the antigenemia assay.Finally, the application of a fluorescent dye detection system and image analysis of the acrylamide gel with a laser scanner provides additional sensitivity to the detection of cytomegalovirus, as well as avoiding the use of radioactivity, making the assay more adaptable to the clinical laboratory.

Congenital cytomegalovirus infection in infants infected with human immunodeficiency virus type 1.

To determine the rate of in utero transmission of cytomegalovirus (CMV) in perinatally HIV-exposed infants and to determine whether coinfection with CMV in early life affects outcome. Infants born to HIV-infected women between March, 1988, and March, 1995, were evaluated (n = 206). Congenital or in utero CMV infection was defined as a positive CMV culture or shell vial assay on urine obtained in the first 3 weeks of life. HIV-infected infants either had positive serology beyond 18 months of age or, for an infant younger than 18 months, had a positive HIV PCR or HIV culture on at least two separate occasions. There were 30 HIV-infected and 171 uninfected infants (144 who seroreverted and 27 infants with at least 2 negative HIV PCR or culture results and normal immunologic studies during the first 6 months of age). Urine culture for CMV was obtained during the first 3 weeks of life on 154 infants: 24 of 30 (80%) HIV-infected infants; and 130 of 171 (76%) HIV-uninfected infants. Overall 10 of 154 (6.5%) infants were infected with CMV: 5 of 24 (21%) HIV-infected and 5 of 130 (3.8%) HIV-uninfected infants. The rate of in utero CMV infection was significantly higher in HIV-infected infants (P = 0.008). Dually infected infants were more immunosuppressed than their CMV-negative counterparts. At 3 months of age the percentage of CD4+ T lymphocytes (P = 0.0021) and CD4:CD8 ratios (P = 0.0018) were significantly lower in the CMV-infected infants than in the CMV-uninfected infants. At 6 months of age the absolute CD4+ T lymphocyte counts (P = 0.0038), percentage of CD4+ T lymphocytes (P = 0.044) and CD4:CD8 ratios (P = 0.037) were significantly lower in the CMV-infected infants. The mean survival of HIV-infected infants who were coinfected with CMV in early life (5 in utero and 1 perinatally infected infant identified at 7 weeks) was 24.77 months. Kaplan-Meier survival analysis indicated a trend toward decreased survival in the infants who were coinfected with CMV in early life (P = 0.088). Congenital CMV infection is more common in HIV-infected infants than in HIV-uninfected infants. Infection with CMV in early life is associated with greater immunosuppression and may be associated with a more rapid progression of HIV infection in infants.